An insulin-related signaling pathway in the nematode Caenorhabditis elegans controls aging and life span. Remarkably, the C. elegans genome contains 38 putative insulin-like genes and a single orthologue of the human insulin receptor (designated daf-2). Do nematode insulin-like proteins truly resemble mammalian insulins in structure and function? Why are there such a diversity of ligands? We propose to synthesize representative insulin-like polypeptides (INS proteins) and investigate their structures and receptor-binding properties. The proposed studies will illuminate the evolution of the insulin structure and dissect a signaling pathway in a genetic model of senescence. The proposed studies focus on four questions. (1) Given their sequence divergence, are INS proteins truly members of the insulin superfamily, and if so, how are the divergent side chains accommodated within a conserved architecture? (2) Does DAF-2 function as a receptor tyrosine kinase, and if so, is its kinase activity positively or negatively regulated by INS ligands? (3) Do INS proteins bind to the human insulin receptor, and if so, how can their seemingly divergent surfaces mimic human insulin? (4) Conversely, does human insulin bind to and regulate DAF-2? A long-term collaboration is proposed with Prof. Guy G. Dodson to compare co-crystal structures of hormone-receptor complexes. Preliminary studies of a representative INS protein have established that -- despite an overall insulin-like fold -- its putative receptor-binding surface differs dramatically from that of human insulin. As a seeming paradox, the protein nonetheless binds to and activates the human insulin receptor. Human insulin likewise binds to the DAF-2 receptor. Novel INS ligands may provide an unexpected starting point for design of human insulin agonists. Given the growing importance of insulin analogs to optimize treatment of diabetes mellitus and the possible clinical utility of IGF-I antagonists in cancer therapy, studies of the INS family may have translational implications in therapeutic protein design.